Abstract
BACKGROUND: In vascular smooth muscle, rises in intracellular calcium [Ca(2+)](i) drive contraction downstream of α(1)-adrenoceptor activation via IP3. Endothelium-derived 6-nitrodopamine (6-ND) augments cardiac catecholamine actions; however, its effect on [Ca(2+)](i) is unknown. We hypothesized that 6-ND would increase intracellular [Ca(2+)](i) and potentiate catecholamine-induced [Ca(2+)](i) signalling in vascular smooth muscle cells, with resultant functional effects on vascular tone when measured in vitro. METHODS: Human aortic smooth muscle cells (HASMCs) were loaded with fura-2 AM (1 μM) and [Ca(2+)](i) measured using a CLARIOstar plate reader after addition of Hanks' balanced salt solution. Rat thoracic aorta rings, with the endothelium removed, were mounted in Krebs-Henseleit baths, and isometric force was recorded via PowerLab. RESULTS: 6-ND and classical catecholamines evoked concentration-dependent increases in HASMC Ca(2+) flux, with 6-ND displaying the greatest potency. 6-ND potentiated the increases in HASMC Ca(2+) flux induced by the classical catecholamines. Tetrodotoxin (TTX) caused a concentration-dependent inhibition of responses to 6-ND and dopamine but did not alter noradrenaline- or adrenaline-induced [Ca(2+)](i) rise. In endothelium-denuded aortic rings, 6-ND potentiated contractions elicited by catecholamines, and this potentiation was abolished by TTX. CONCLUSIONS: 6-ND is a potent modulator of [Ca(2+)](i) in HASMCs and enhances catecholamine-driven vasoconstriction. Both effects are blocked by TTX, indicating that 6-ND modulates voltage-gated sodium channels upstream of Ca(2+) entry/release in vascular smooth muscle cells.